Spatially Defined Cell-Secreted Protein Detection Using Granular Hydrogels: μGeLISA

Abstract

Intercellular communication through secreted proteins is necessary in essential processes such as embryo and limb development, disease progression, and immune responses. There exist many techniques to study bulk solution protein concentrations, but there is a limited set of tools to study the concentrations of cell-secreted proteins in situ within diverse cell platforms while retaining spatial information. In this study, we have developed a microgel system that is able to quantitatively measure the cell-secreted protein concentration within defined threedimensional culture configurations with single-cell spatial resolution, called μGeLISA (microgel-linked immunosorbent assay). This system, which is based on the surface modification of polyethylene glycol microgels, was able to detect interleukin 6 (IL-6) concentrations of 2.21−21.86 ng/mL. Microgels were also able to detect cell spheroid-secreted IL-6 and distinguish between lowand high-secreting single cells. The system was also adapted to measure the concentration of cell-secreted matrix metalloproteinase-2 (MMP-2). μGeLISA represents a highly versatile system with a straightforward fabrication process that can be adapted toward the detection of secreted proteins within a diverse range of cell culture configurations.

Combinatorial Microgels for 3D ECM Screening and Heterogeneous Microenvironmental Culture of Primary Human Hepatic Stellate Cells

“…The 24% dextran and 16% MgSO4 solution was added to this PEG solution, creating an aqueous two‐phase system that can be vortexed to form a microdroplet suspension of PEG within the dextran bath. [ 41 , 42 ] The suspension was irradiated with UV light for 90 s to allow for the microdroplets to polymerize and form microgels ( Figure 1 a ). Interestingly, when the protein was covalently crosslinked to the PEG backbone at the same time as the polymerization, the protein was distributed in the core or on the outer edge of the µG.…”

Advances in high throughput cell culture technologies for therapeutic screening and biological discovery applications

“…Often also called granular hydrogel scaffolds, microscale hydrogel scaffolds are functionalized with bioadhesive ligands to provide a microporous scaffold for 3D cell culture upon assembly (Figure 4f ). 207 The most common methods of fabricating microgels for suitability as a cell culture scaffold are mechanical fragmentation, 208 , 209 batch emulsion, 210 , 211 microfluidics 212 , 213 and photolithography. 214 , 215 In contrast to the cell‐encapsulated microgels mentioned previously, these microgels can be made in harsher manufacturing environments as cell viability during polymerization or formation of droplets is not a design consideration, due to the timing of cell introduction which occurs subsequent to microgel fabrication.…”